Quorum sensing is a bacterial cell-to-cell signaling system in which communication occurs through hormone like organic compounds referred to as auto-inducers. These auto-inducers then interact with bacterial transcription factors to drive gene expression in a coordinate manner. Quorum sensing has also recently been recognized as a cell-to-cell signaling system amongst prokarytes and eukaryotes. In particular, the AI-3 bacterial signaling system cross-signals with the human epinephrine/norepinephrine signaling system. The AI-3/epinephrine/norepinephrine signaling cascade is present in several bacterial pathogens such as enterohemorrhagic E. coli (EHEC) O157:H7, Salmonella, Shigella, Yersinia, Francisella tularensis, among others. We have extensivelly demonstrated that this signaling system is responsible for activating EHEC virulence genes. Given the widespread nature of this signaling system amongst several bacterial pathogens, and its defined role in EHEC pathogenesis, this proposal aims to identify molecular chemical inhibitors of this quorum sensing system using a high throughput screen (HTS). This screen will use the UT Southwestern chemical compound library to identify inhibitors of AI-3 signaling. We already have preliminary data on primary screens that were performed and Specific Aim 1 of this proposal is designed to validate and confirm these AI-3 inhibitors. Specific Aim 2 will expand the utilization of these inhibitors in several virulence models of EHEC virulence. Given the multi-drug resistance of several bacterial pathogens to conventional antibiotics, and the need to develop novel classes of antimicrobials, these studies may help to generate a whole new class of antimicrobials that can block AI-3 and epinephrine signaling to bacterial pathogens.